Anti-SUMO2/3 antibody is a SUMO-2/3 mouse monoclonal antibody that is part of the Signal-Seeker™ product line. Anti-SUMO-2/3 mouse monoclonal antibody was raised against full-length recombinant SUMO-2 protein (Uniprot: P61956) combined with a proprietary mix of peptides that include CQIRFRFDGQPINE. The antibody has been shown to recognize a wide range of SUMO-2/3-targeted proteins in HeLa cell lysate (Fig. 1B) and to detect sub-nanogram amounts of recombinant SUMO-2 (Fig. 1A). Epitope mapping has identified that the antibody recognizes a sequence/structure within the peptide CQIRFRFDGQPINE. The peptide sequence is conserved in mammals, birds, and amphibians, giving the antibody broad species reactivity. ASM23 is purified by Protein G affinity chromatography and is supplied as a lyophilized white powder.

Each Lot of antibody is quality controlled to provide a high batch to batch consistency. The Lot specific µg per tube can be found in the Lot specific COA documents.

Denatured cell lysates were prepared from HS43, CT37 and KD S212. 1mg of lysate was used for the immunoprecipitation of SUMO-2/3 conjugates. IP experiments were performed by the protocol presented in IP Method. Western blots of immunoprecipitated proteins were developed using 12F3 (A) or anti-TFII-I antibody (B). (A) Star (*) and circle (o) indicate heavy and light chains of antibodies. Unconjugated free SUMO is denoted by triangle. (B) Unconjugated TFII-I is visible near 120kDa. Multiple bands indicate that TFII-I is SUMOylated by several SUMO-2/3 proteins. TFII-I has previously been reported to be a target for Sumoylation . To see the full Immunoprecipitation protocol, see the product datasheet.

HeLa cells were stained and visualized by confocal fluorescence microscopy as described in the IF method below. The cells were stained against α/β-tubulin (sheep anti-tubulin Ab, Cat# ATN02, green) and SUMO-2/3 (12F3, red). DNA was stained with DAPI. Mitotic cells in metaphase were imaged with a Zeiss LSM 780 confocal microscope (1.4 NA 63X objective). Detection of SUMO 2/3 at chromosomes can be observed during mitosis as has been previously reported10. To see the full Immunofluorescence protocol, see the product datasheet.

HeLa cells were stained and visualized by widefield fluorescence microscopy as described in the IF method below. The cells were stained against α/β-tubulin (sheep anti-tubulin Ab, Cat# ATN02, green) and SUMO-2/3 (12F3, red). DNA was stained with DAPI. Cells in interphase were imaged with a Zeiss Axio Observer.Z1 microscope (1.4 NA 63X objective). PML nuclear bodies (nuclear dots) were visible in SUMO-2/3 staining as has been previously reported11. To see the full Immunofluorescence protocol, see the product datasheet.